In many instances it is desirable to grow large numbers of genetically identical plants. These plants can be selected and grown based on their particular qualities, such as their ability to grow in a particular climate, or their ability to produce a particular type or quality of fiber. Unfortunately, in many cases the production of such plants through standard breeding is not feasible.
Standard plant breeding techniques are labor intensive and, because they usually involve fusion of gametes, tend to introduce genetic variability. The genetic variability is a result of crossing over of chromosomes, which often occurs during meiosis. Additionally, standard breeding techniques require that the breeder wait until a plant is mature enough to breed. In some cases, this means waiting many years. This delay decreases productivity and increases costs. Therefore, it is desirable to produce large numbers of genetically identical plants via culturing of somatic or zygotic plant embryos.
Somatic or zygotic plant embryos may be cultured in the form of “manufactured seeds”. Manufactured seeds are essentially analogs of botanic seed and typically include a nutrient medium (termed here in a “nutritive medium”), and typically include structural features that serve to protect the embryo before, during, and after germination. References that describe manufactured seeds include U.S. Pat. Nos. 5,564,224; 5,687,504; and 5,701,699, all to Carlson et al. These patents disclose elements of such seeds, for instance, a manufactured seed coat, methods for using such manufactured seeds, and plant germinants produced from manufactured seeds.
A problem with manufactured seeds to date is the relatively low numbers of successful germinants from such seeds compared to botanical (natural) seeds, and relatively low numbers of “normal” germinants from manufactured seeds, i.e., viable, uniform, and commercially useful germinants. Abnormal germinants possess any of various malformations such as small and/or deformed radicals, hypocotyls, or cotyledons. Although many factors appear to cause abnormal germination, the results generally indicate that manufactured seeds, as currently known in the art, do not provide a correct balance of nutrients and other elements to the seed.
Therefore, there is a need for nutritive media that contain a suitable profile of nutrients and other elements that, when incorporated into manufactured seeds, facilitate normal and successful germination and germinant growth with normal organ development.
In order to be commercially viable, a nutritive medium desirably facilitates the germination of manufactured seeds at a rate at least similar to the germination rate of sexually produced seeds. Commercial viability can be assessed by taking into account the market size and the volume of product necessary to meet market needs. In the case of plant-related products the market is vast. Therefore, a difference in the germination rate of 10% or 20% can have an immense impact on the commercial viability of manufactured seeds.
Researchers have investigated the effects caused by the addition of various compounds to nutritive media in order to further the understanding of germination and/or growth of plant embryos on such media. For example, the effect of various nutrients on chlorophyll formation in embryos grown in the dark has been studied by Bogorad, Botanical Gazette 3: 221-241, 1950, and by Engvild, Physiologia Plantarum 17: 866-874, 1964; the effect of sucrose concentration has been studied by Schau, Physiologia Plantarum 4: 617-620, 1951, and by Ball, American Journal of Botany 46: 130-139, 1958; the effect of cytokinins on germination has been studied by Khan, Science 71:853-859, 1971; the effect of smoke on germination has been studied by Brown and Van Staden, Plant Growth Regulation 22:115-124, 1997; and the effect of amino acids as sources of organic nitrogen has been studied by Engvild (cited above).
In the laboratory, growth of plant embryos on the surface of an agar medium in a petri dish or the like is known. Unfortunately, the correlation between the germination of a plant embryo on the surface of an agar medium and the germination of an embryo from a manufactured seed is weak. This is because inter alia, there are many substantial differences between bare plant embryos and manufactured seeds. A key difference is the physical structure of the seed, which can have a profound impact on survival of the embryo inside the seed and on germination success.
There also remain large differences between manufactured seeds and corresponding natural seeds. Whereas, the embryo relies on the megagametophyte for nutrients useful for germination, the embryo in a manufactured seed relies on the nutritive medium that is provided in the manufactured seed.
Also, an agar medium used as a surface on which plant embryos are grown appears to have substantially different requirements compared to a nutritive medium for use in manufactured seed.
Therefore, there is a need for nutritive media that, when incorporated into manufactured seeds, provides an improved germination rate compared to conventional manufactured seeds.